The resistance of pathogens to various antimicrobial agents has increased at an alarming rate in recent years rendering many important therapeutics for the treatment of microbial infections ineffective. There exist multiple mechanisms for resistance that have been well studied. Pathogens may employ one or more modes of resistance rendering them polyresistant. Polyresistant pathogens are found among a variety of microorganisms including rendering some treatable by only a single class of clinically available antimicrobial agents, if at all.
In response to the rapid development of polyresistant pathogens, combinations of active antimicrobial therapeutic agents are being employed to treat disease. The most useful combination therapies appear to be capable of inhibiting or killing microbes via multiple mechanisms thereby circumventing microbial resistance.
Thus there is a need for antimicrobial agents or a combination of agents that together possess potent broad spectrum antimicrobial activity to which the microbe can not easily become resistant, and can be administered at lower concentrations while maintaining or improving therapeutic activity, and have reduced toxicity, and are inexpensive to produce.
In one aspect, the present invention relates to antimicrobial compositions.
Another aspect of the invention deals with pharmaceutical compositions comprising a broad group of known antibacterial agents in combination with polymers of the invention as defined herein and a pharmaceutically acceptable carrier.
Yet another aspect of the invention is a treatment regimen (also referred to herein as the xe2x80x9ctreatment regimenxe2x80x9d or xe2x80x9cmethodxe2x80x9d of the invention) for treating a microbial infection in a mammal, comprising administering to the mammal a therapeutically effective amount of polymer as defined herein in combination with a therapeutically effective amount of an antibacterial agent.
The polymer to be administered can be a homopolymer or a copolymer. In one embodiment, the polymer further includes a monomer comprising a hydrophobic group, such as an aryl group or a normal or branched C3-C18-alkyl group.
The polymer to be administered can, optionally, further include a monomer comprising a neutral hydrophilic group, such as a hydroxyl group or an amide group.
The polymer can further have a backbone which is interrupted at one or more points by a nitrogen containing functional group such as a quaternary ammonium group, phosphorus containing functional groups, or sulfur containing functional groups.
Preferred polymers of the invention include amine or ammonium functional groups attached to the polymer backbone via aliphatic spacer groups.
The term xe2x80x9cantimicrobial agentsxe2x80x9d are intended to include antibacterial agents, antifungal agents, antiseptics and the like.
The treatment regimen of the invention represents a new approach to antibacterial therapy to which a polymer can be administered in combination with an antibacterial agent to provide a therapeutically effective treatment of an infection, particularly an infection involving resistant or polyresistant bacteria, and/or to reduce the overall amount of antibacterial agent, or polymer necessary to treat an infection.
Furthermore, the treatment regimen of the invention may reduce the need to develop new antibacterial agents as bacteria and other microbes develop resistance to known antibacterial and antimicrobial agents.
In addition, the polymers employed in the invention are easily prepared using standard techniques of polymer synthesis and inexpensive starting materials. Preferably, the polymers will not be substantially degraded in the digestive tract and, therefore, can be administered orally or topically. Polymer compositions can also be readily varied, to optimize properties such as solubility or water swellability and antimicrobial activity.
A description of preferred embodiments of the invention follows.
The present invention relates to antimicrobial compositions including pharmaceutical compositions and treatment regimens for preventing or treating a microbial infection in a mammal, such as a human, by administering to the mammal, a therapeutically effective amount of an cationic polymer and a therapeutically effective amount of an antibacterial agent.
As used herein, a xe2x80x9ctherapeutically effective amountxe2x80x9d is a first amount of a polymer in combination with a second amount of an antimicrobial agent that is sufficient to therapeutically inhibit, partially or totally, a microbial infection, or to reverse development of a microbial infection, or prevent or reduce its further progression.
The term xe2x80x9cantimicrobial agentxe2x80x9d is intended to include antibacterial agents, antifungal agents, antiseptics and the like. The term xe2x80x9cantibacterial agentxe2x80x9d includes but is not limited to: naturally occurring antibiotics produced by microorganisms to suppress the growth of other microorganisms, and agents synthesized or modified in the laboratory which have either bacteriocidal or bacteriostatic activity, e.g., xcex2-lactam antibacterial agents including, e.g. carbencillim; ampicillin, cloxacillin, oxacillin and pieracillin, cephalosporins and other cephems including, e.g. cefaclor, cefamandole, cefazolin, cefoperazone, ceftaxime, cefoxitin, ceftazidime, ceftriazone and carbapenems including, e.g., imipenem and meropenem; and glycopeptides, macrolides, quinolones (e.g. nalidixic acid), tetracyclines, aminoglycosides (e.g. Gentamicin and Paromomycin) and further includes antifungal agents. In general if an antibacterial agent is bacteriostatic, it means that the agent essentially arrests or inhibits bacterial cell growth (but does not kill the bacteria); if the agent is bacteriocidal, it means that the agent kills bacterial cells (and may stop growth before killing the bacteria).
The term xe2x80x9cpolymerxe2x80x9d refers to a macromolecule comprising a plurality of repeat units or monomers. The term includes homopolymers, which are formed from a single type of monomer, and copolymers, which are formed of two or more different monomers. A xe2x80x9cterpolymerxe2x80x9d is a copolymer formed from three different monomers. The term polymer, as used herein, is intended to exclude proteins, peptides, polypeptides and proteinaceous materials.
As used herein, the term xe2x80x9cpolymer backbonexe2x80x9d or xe2x80x9cbackbonexe2x80x9d refers to that portion of the polymer which is a continuous chain, comprising the bonds which are formed between monomers upon polymerization. The composition of the polymer backbone can be described in terms of the identity of the monomers from which it is formed, without regard to the composition of branches, or side chains, off of the polymer backbone. Thus, a poly(acrylamide) polymer is said to have a poly(acrylamide) backbone, without regard to the substituents on the acrylamide nitrogen atom, which are components of the polymer side chains. A poly(acrylamide-co-styrene) copolymer, for example, is said to have a mixed acrylamide/styrene backbone.
The term xe2x80x9cpolymer side chainxe2x80x9d or xe2x80x9cside chainxe2x80x9d refers to the portion of a monomer which, following polymerization, forms a branch off of the polymer backbone. In a homopolymer all of the polymer side chains are identical. A copolymer can comprise two or more distinct side chains. When a side chain comprises an ionic unit, for example, the ionic unit depends from, or is a substituent of, the polymer backbone, and is referred to as a xe2x80x9cpendant ionic unitxe2x80x9d. The term xe2x80x9cspacer groupxe2x80x9d, as used herein, refers to a polyvalent molecular fragment which is a component of a polymer side chain and connects a pendant moiety to the polymer backbone. The term xe2x80x9caliphatic spacer groupxe2x80x9d refers to a spacer group which does not include an aromatic unit, such as a phenylene unit.
The term xe2x80x9caddition polymerxe2x80x9d, as used herein, is a polymer formed by the addition of monomers without the consequent release of a small molecule. A common type of addition polymer is formed by polymerizing olefinic monomers, wherein monomers are joined by the formation of a carbon-carbon bonds between monomers, without the loss of any atoms which compose the unreacted monomers.
The term xe2x80x9cmonomerxe2x80x9d, as used herein, refers to both (a) a single molecule comprising one or more polymerizable functional groups prior to or following polymerization, and (b) a repeat unit of a polymer. An unpolymerized monomer capable of addition polymerization, can, for example, comprise an olefinic bond which is lost upon polymerization.
For the treatment regimen of the invention, the appropriate combination of polymer and antibiotic to be administered for treatment of a microbial infection in a mammal will be determined on an individual basis and will be determined, at least in part, by consideration of the individual""s size, the severity of symptoms to be treated and the result sought.
The treatment regimen encompasses co-administration of therapeutically effective amounts of polymer and antibacterial agent in a single, substantially simultaneous manner, such as in a single capsule, tablet, injection or topical ointment having a fixed ratio of polymer and antibacterial agent, or in multiple, separate capsules, tablets, ointments or injections for each of the polymer and antibacterial agent. In addition, the treatment regimen also encompasses use of each compound separately, in a sequential manner (e.g. minutes or hours apart).
Each component of the treatment regimen may be administered, for example, topically, orally, intranasally, or rectally. The form in which the agent is administered, for example, powder, tablet, capsule, solution, or emulsion, depends in part on the route by which it is administered. The therapeutically effective amount of each component of the regimen can be administered in a series of doses separated by appropriate time intervals, such as hours.
Microbial infections which can be treated or prevented by the treatment regimen of the invention include bacterial protozoal, viral, ameobic, fungal and parasitic infections, such as infection by Streptococcus, including Streptococcus mutans, Streptococcus salivarius, and Streptococcus sanguis, Salmonella, Campylobacter, including Campylobacter sputum and Campylobacter jejuni, Heliobacter, including Heliobacter pylori, Antinomyces, including Actinomyces naeslundii and Actinomyces viscosus, Escherichia coli, Clostridium difficile, Staphylococcus, including S. aureus, Shigella, Pseudomonas, including P. aeruginosa, Eikenella corrodens, Actinobacillus actinomycetemcomitans, Bacteroides gingivalis, Capnocytophaga, including Capnocytophaga gingivalis, Wolinell recta, Bacteriodes intermedius, Mycoplasma, including Mycoplasma salivarium, Treponema, including Treponema denticola, Peptostreptococcus micros, Bacteriodes forsythus, Fusobacteria, including Fusobacterium nucleatum, Selenomonas sputigena, Bacteriodes fragilis, Enterobacter cloacae, Pneumocystis, Cryptosporidium parvumand Giardia lamblia, Entameoba histolytica or Acanthameoba, such as A. castellani, Candida albicans, Aspergillus fumigatus, and Trichinella spiralis. The method is useful for treating infections of various organs of the body, but is particularly useful for infections of the skin and gastrointestinal tract.
Suitable polymers for the present treatment method include polymers having amine or ammonium functional groups attached to the polymer backbone via aliphatic spacer groups.
Polymers which are particularly suitable for the present method include polymers that form amphipathic structures. The term xe2x80x9camphipathicxe2x80x9d, as used herein, describes a three-dimensional structure having discrete hydrophobic and hydrophilic regions. Thus, one portion of the structure interacts favorably with aqueous and other polar media while another portion of the structure interacts favorably with non-polar media. An amphipathic polymer results from the presence of both hydrophilic and hydrophobic structural elements along the polymer backbone. 
wherein X is a covalent bond, a carbonyl group or a CH2 group, Y is an oxygen atom, an NH group or a CH2 group, Z is a spacer group, R is a hydrogen atom or a methyl or ethyl group; R1, R2 and R3 are each, independently, a hydrogen atom, a normal or branched, substituted or unsubstituted C1-C24-alkyl group, an aryl group or an arylalkyl group; Axe2x88x92 is a pharmaceutically acceptable anion, such as a conjugate base of a pharmaceutically acceptable acid; and m and n are each, independently, 0 or 1. Suitable alkyl substituents include halogen atoms, such as fluorine or chlorine atoms. A monomer of Formula 1 in which at least one of substituents R1, R2 and R3 is hydrogen can also exist in the free base, or amino, form in which a hydrogen substituent is absent and the nitrogen atom is electrically neutral.
In a preferred embodiment, one of R1-R3 is an ammonioalkyl group of the general formula 
wherein R4, R5 and R6 are each, independently, a hydrogen atom, a C1-C24 alkyl group, or an arylalkyl group; n is an integer from 2 to about 20, preferably from 3 to about 6; and Axe2x88x92 is a pharmaceutically acceptable anion. An ammonioalkyl group in which at least one of substituents R4, R5 and R6 is hydrogen can also exist in the free base, or amino, form in which a hydrogen substituent is absent and the nitrogen atom is electrically neutral. The group xe2x80x94N+(R4)(R5)(R6) can also be a heteroaryl group, such as a 5- or 6-membered heteroaryl group, such as a 1-pyridinio group. Preferably, at least one of R4, R5 and R6 is a C6-C24-alkyl group. Examples of suitable ammonioalkyl groups include, but are not limited to, 4-(dioctylmethylammonio)butyl; 3-(dodecyldimethylammonio)propyl; 3-(octyldimethylammonio)propyl; 3-(decyldimethylammonio)propyl; 5-(dodecyldimethylammonio)pentyl; 3-(cyclohexyldimethylammonio)propyl; 3-(decyldimethylammonio)-2-hydroxypropyl; 3-(tridecylammonio)propyl; 3-(docosyldimethylammonio)propyl; 4-(dodecyldimethylammonio)butyl; 3-(octadecyldimethylammonio)propyl; 3-(hexyldimethylammonio)propyl; 3-(methyldioctylammonio)propyl; 3 -(didecylmethylammonio)propyl; 3-(heptyldimethylammonio)propyl; 3-(dimethylnonylammonio)propyl; 6-(dimethylundecylammonio)hexyl; 4-(heptyldimethylammonio)butyl; 3-(dimethylundecylammonio)propyl; 3-(tetradecyldimethylammonio)propyl; 3-(1-pyridinium)propyl; in combination with a pharmaceutically acceptable anion.
When at least one of R1 to R6 is a hydrogen atom, the monomer can also exist in the free base, or amino form. The polymer comprising such a monomer can be administered in the free base form or in the protonated or partially protonated form, for example, as a salt of a pharmaceutically acceptable acid. Suitable acids include hydrochloric acid, hydrobromic acid, citric acid, lactic acid, tartaric acid, phosphoric acid, methanesulfonic acid, acetic acid, formic acid, maleic acid, fumaric acid, malic acid, succinic acid, malonic acid, sulfuric acid, L-glutamic acid, L-aspartic acid, pyruvic acid, mucic acid, benzoic acid, glucoronic acid, oxalic acid, ascorbic acid, and acetylglycine. In either case, at physiological pH following administration, a plurality of amino groups will be protonated to become ammonium groups, and the polymer will carry an overall positive charge.
The spacer group Z is a component of the polymer side chain and connects the amino or ammonium group to the polymer backbone. The amino or ammonium group is, thus, a pendant group. The spacer group can be a normal or branched, saturated or unsaturated, substituted or unsubstituted alkylene group, such as a polymethylene group xe2x80x94(CH2)nxe2x80x94, wherein n is an integer from about 2 to about 24. Suitable examples include the propylene, hexylene and octylene groups. The alkylene group can also, optionally, be interrupted at one or more points by a heteroatom, such as an oxygen, nitrogen (e.g, NH) or sulfur atom. Examples include the oxaalkylene groups xe2x80x94(CH2)2O[(CH2)2O]n(CH2)2xe2x80x94, wherein n is an integer ranging from 0 to about 3.
Examples of monomers of Formula I having quaternary ammonium groups include:
N-(3-dimethylaminopropyl)acrylamide,
N-(3-trimethylammoniopropyl)acrylamide,
2-trimethylammonioethyl methacrylate,
2-trimethylammonioethyl acrylate,
N-(3-trimethylammoniopropyl)methacrylamide,
N-(6-trimethylammoniohexyl)acrylamide,
N-(3-trimethylammoniopropyl)acrylamide,
N-(4-trimethylammoniobutyl)allylamine,
N-(3-dimethyloctylammoniopropyl)allylamine,
N-(3-trimethylammoniopropyl)allylamine,
N-(3-(1-pyridinio)propyl)vinylamine and
N-(3-(1-pyridinio)propyl)allylamine.
Each of these monomers also includes a suitable counter anion. Examples of monomers of Formula I having an amino group include allylamine, vinylamine and N-(3-dimethylaminopropyl)acrylamide. Each of these monomers can also exist as a salt with a pharmaceutically acceptable acid.
In another embodiment, the polymer to be administered is characterized by a diallylamine repeat unit of Formula III: 
wherein R1 and R2 are each, independently, a hydrogen atom, a normal or branched, substituted or unsubstituted C1-C24-alkyl group, an aryl group or an arylalkyl group; and Axe2x88x92 is a pharmaceutically acceptable anion, such as a conjugate base of a pharmaceutically acceptable acid. Suitable alkyl substituents include halogen atoms, such as fluorine or chlorine atoms. A monomer of Formula III in which at least one of substituents R1 and R2 is hydrogen can also exist in the free base, or amino, form, in which a hydrogen substituent is absent and the nitrogen atom is electrically neutral. In a preferred embodiment, R1 is an ammonioalkyl group of Formula II, as described above.
In another embodiment, the polymer to be administered is a poly(alkyleneimine) polymer comprising a monomer, or repeat unit, of Formula IV, 
wherein n is an integer from about 2 to about 10 and R7 and R8 are each, independently, a hydrogen atom, a normal or branched, substituted or unsubstituted C1-C24-alkyl group, an aryl group or an arylalkyl group, and Axe2x88x92 is a pharmaceutically acceptable anion. Suitable alkyl substituents include halogen atoms, such as fluorine or chlorine atoms. When one of R7 and R8 is a hydrogen atom, the polymer can be administered in the free base form or in the cationic form shown, as the salt of a pharmaceutically acceptable acid. A monomer of Formula IV in which at least one of substituents R7 and R8 is hydrogen can also exist in the free base, or amino form, in which a hydrogen substituent is absent and the nitrogen atom is electrically neutral. In a preferred embodiment, the polymer to be administered is a poly(ethyleneimine) polymer, comprising a monomer of Formula IV wherein n is 2.
Preferably, R7 is an aminoalkyl group, or an ammonioalkyl group of Formula II, as described above. In one embodiment, the polymer comprises monomeric units of Formula 11 wherein R7 is an aminoalkyl group, or an ammonioalkyl group, as well as monomeric units wherein R7 and R8 are each hydrogen or R7 is hydrogen and R8 is absent. The fraction of monomeric units which include the aminoalkyl or ammonioalkyl group can be from about 5% to about 90% of the monomeric units of the polymer.
The spacer group is a component of the polymer side chain and connects the amino or ammonium group to the polymer backbone. The amino or ammonium group is, thus, a pendant group. The spacer group can be a normal or branched, saturated or unsaturated, substituted or unsubstituted alkylene group, such as a polymethylene group xe2x80x94(CH2)nxe2x80x94, wherein n is an integer from about 2 to about 15. Suitable examples include the propylene, hexylene and octylene groups. The alkylene group can also, optionally, be interrupted at one or more points by a heteroatom, such as an oxygen, nitrogen (e.g, NH) or sulfur atom. Examples include the oxaalkylene groups xe2x80x94(CH2)2O[(CH2)2O]n(CH2)2xe2x80x94, wherein n is an integer ranging from 0 to about 3.
Polymers to be administered which have quaternary ammonium groups or protonated amino groups will further comprise a pharmaceutically acceptable counter anion, such as anions which are conjugate bases of the pharmaceutically acceptable acids discussed above, for example, chloride, bromide, acetate, formate, citrate, ascorbate, sulfate or phosphate. The number of counter anions associated with the polymer prior to administration is the number necessary to balance the electrical charge on the polymer.
The polymer can also be a copolymer further comprising a hydrophobic monomer. The hydrophobic monomer can comprise a side chain bearing a hydrophobic group, such as a straight chain or branched, substituted or unsubstituted C3-C18-alkyl group or a substituted or unsubstituted aryl group. Examples of suitable hydrophobic monomers include styrene, N-isopropylacrylamide, N-t-butylacrylamide, N-n-butylacrylamide, heptafluorobutylacrylate, N-n-decylallylamine, N-n-decylacrylamide, pentafluorostyrene, n-butylacrylate, t-butylacrylate, n-decylacrylate, N-t-butylmethacrylamide, n-decylmethacrylate, and n-butylmethacrylate.
Examples of copolymers comprising a monomer of Formula I and a hydrophobic monomer include poly(N-(3-dimethylaminopropyl)acrylamide-co-N-(n-butyl)acrylamide) or salts thereof with pharmaceutically acceptable acids. Other examples of suitable copolymers include poly(2-trimethylammoniumethylmethacrylate-co-styrene) chloride, poly(2-trimethylammoniumethylmethacrylate-co-N-isopropylacrylamide) chloride, poly(2-trimethyl-ammoniumethylmethacrylate-co-heptafluorobutylacryl) chloride, poly(3-trimethylammoniumpropylmethacrylate-co-styrene) chloride, poly(3-trimethylammoniumpropylmethacrylate-co-N-t-butylacrylamide) chloride, poly(3-trimethylammoniumpropylmethacrylate-co-N-n-butylacrylamide) chloride, and poly(N-(3-trimethylammoniumpropyl)allylamine-co-N-n-decylallylamine). Each of these ionic copolymers can also be employed with counter ions other than chloride, for example, a conjugate base of a pharmaceutically acceptable acid.
In a further embodiment, the polymer to be administered comprises a monomer of Formula I, a hydrophobic monomer and a neutral hydrophilic monomer, such as acrylamide, methacrylamide, N-(2-hydroxyethyl)acrylamide or 2-hydroxyethylmethacrylate. Examples of polymers of this type include terpolymers of N-(3-trimethylammonium-propyl)methacrylamide/N-isopropylacrylamide/2-hydroxyethyl-methacrylate, N-(3-trimethylammonium-propyl)methacrylamide/N-n-decylacrylamide/2-hydroxyethylmethacrylate, N-(3-trimethylammoniumpropyl)methacrylamide/N-t-butylmethacrylamide/methacrylamide, N-(3-trimethylammoniumpropyl) methacrylamide/n-decylacrylate/methacrylamide, 2-trimethylammoniumethylmeth-acrylate/n-butyl-acrylate/acrylamide, 2-trimethylammonium-ethylmethacrylate/t-butylacrylate/acrylamide, 2-trimethylammoniumethylmethacrylate/n-decylacrylate/acrylamide, 2-trimethylammonium-ethylmethacrylate/n-decylmethacrylate/methacrylamide, 2-trimethylammoniumethylmethacrylate/N-t-butyl-methacrylamide/methacrylamide and 2-trimethylammoniumethylmethacrylate/N-n-butyl-methacrylamide/methacrylamide.
The polymer to be administered can be an addition polymer having a polymer backbone such as a polyacrylate, polyacrylamide, poly(allylalcohol), poly(vinylalcohol), poly(vinylamine), poly(allylamine), or polyalkyleneimine backbone. The polymer can have a uniform backbone if it is composed of monomers derived from a common polymerizable unit, such as acrylamide. If the polymer is a copolymer, it can also comprise a mixed backbone, for example, the monomer of Formula I can be an acrylamide derivative, while the hydrophobic monomer can be a styrene derivative. The polymers disclosed herein include examples of both uniform and mixed backbones.
The polymers of use in the present method also include condensation polymers, wherein polymerization of monomers is accompanied by the release of a small molecule, such as a water molecule. Such polymers include, for example, polyesters and polyurethanes.
The polymers of use in the present method are preferably substantially nonbiodegradable and nonabsorbable. That is, the polymers do not substantially break down under physiological conditions into fragments which are absorbable by body tissues. The polymers preferably have a nonhydrolyzable backbone, which is substantially inert under conditions encountered in the target region of the body, such as the gastrointestinal tract.
The composition of the copolymers to be administered can vary substantially. The copolymer can comprise from about 95 mole percent to about 5 mole percent, preferably from about 20 mole percent to about 80 mole percent, of a monomer of Formula I. The copolymer can also comprise from about 95 mole percent to about 5 mole percent, preferably from about 20 mole percent to about 80 mole percent, of a hydrophobic monomer.
The polymer to be administered will, preferably, be of a molecular weight which is suitable for the intended mode of administration and allows the polymer to reach and remain within the targeted region of the body for a period of time sufficient to interact with the infecting organism. For example, a method for treating an intestinal infection should utilize a polymer of sufficiently high molecular weight to resist absorption, partially or completely, from the gastrointestinal tract into other parts of the body. The polymers can have molecular weights ranging from about 500 Daltons to about 500,000 or to about 1 million Daltons, preferably from about 2,000 Daltons to about 150,000 Daltons, to about 500,000 or to about 1 million Daltons.
The polymers which are useful in the present method can be prepared by known methods. A first method includes the direct polymerization of a monomer, such as trimethylammoniummethylacrylate chloride, or a set of two or more monomers, such as trimethylammoniummethyl-acrylate chloride, N-n-butylacrylamide and acrylamide. This can be accomplished via standard methods of free radical, cationic or anionic polymerization which are well known in the art. Due to reactivity differences between two monomers, the composition of a copolymer produced in this way can differ from the composition of the starting mixture. This reactivity difference can also result in a nonrandom distribution of monomers along the polymer chain.
A second method proceeds via the intermediacy of an activated polymer comprising labile side chains which are readily substituted by a desired side chain. An example of a suitable activated polymer is the succinimide ester of polyacrylic acid, poly(N-acryloyloxysuccinimide) (also referred to hereinafter as xe2x80x9cpNASxe2x80x9d), which reacts with nucleophiles such as a primary amine to form a N-substituted polyacrylamide. Another suitable activated polymer is poly(para-nitrophenylacrylate), which react with amine nucleophiles in a similar fashion.
Polymers suitable for use in the present method can also be prepared by addition of a side chain to a preformed polymer. For example, poly(allylamine) can be alkylated at the amino nitrogen by one or more alkylating agents. For example, one fraction of amino groups can be alkylated using a normal or branched (C3-C18-alkyl halide, such as n-decyl bromide, while another fraction can be alkylate by a quaternary ammonium-containing alkyl halide, such as 1-trimethylammonium-4-bromombutane.
A copolymer having a polyacrylamide backbone comprising amide nitrogens bearing two different substituents can be prepared by treating p(NAS) with less than one equivalent (relative to N-acryloyloxysuccinimide monomer) of a first primary amine, producing a poly(N-substituted acrylamide-co-N-acryoyloxysuccinimide) copolymer. Remaining N-acryoyloxysuccinimide monomer can then be reacted with, for example, an excess of a second primary amine to produce a polyacrylamide copolymer having two different N-substituents. A variety of copolymer compositions can, thus, be obtained by treating the activated polymer with different proportions of two or more amines.
An additional aspect of the present invention is a method for treating a microbial infection in a mammal, such as a human, comprising administering to the mammal a synergistically effective combination therapy regimen comprising an antibacterial agent and a polymer having an amino group or an ammonium group within the polymer backbone. The polymer can have, for example, a polymethylene, backbone which is interrupted by one or more amino or ammonium groups. An example of a polymer of this type is poly(decamethylenedimethylammonium-co-ethylenedimethylammonium) bromide, which is synthesized via the reaction of N,N,Nxe2x80x2,Nxe2x80x2-tetramethylethylenediamine and 1,10-dibromodecane. The polymer can also be administered in association with anions other than bromide, such as chloride or acetate anions. Other examples include poly(alkyleneimines), for example, poly(ethyleneimine). Such polymers can comprise secondary or tertiary amino groups, salts of such groups with pharmaceutically acceptable acids, and/or quaternary ammonium groups.
Examples of other such suitable polymer include polymers comprising piperidine or pyridine groups within the backbone of the polymer (e.g. 4, 4xe2x80x2-Trimethylenedipyridine-alt-1,5 dibromopentane and 4,4-trimethylenepiperidine-alt-a, 6 dibromohexane).
As discussed below in Examples 35-42, several polymers described herein were tested in combination with an antibiotic for in vitro activity against Cryptosporidium parvum infectivity in mammalian cell culture and in vivo activity in mice.
The invention will now be further and specifically described by the following examples.